In recent years, renal cell carcinoma (RCC) has accounted for over 31,000 new cases of cancer and contributed to approximately 12,000 deaths in the United States (Jemal et al. (2002) “Cancer statistics, 2002,” CA—Cancer J. Clin. 52:23-47). Clear cell carcinoma was the predominant subtype comprising up to 85% of RCCs. One-third of patients diagnosed with kidney cancer have evidence of metastatic disease at the time of diagnosis and up to half of those treated for localized disease eventually relapse (Figlin (1999) “Renal cell carcinoma: management of advanced disease,” J. Urol. 161:381-386; discussion 386-387). The natural history of RCC is complex and influenced by factors other than stage (Pantuck et al. (2001) “The changing natural history of renal cell carcinoma,” J. Urol. 166:1611-1623). Patient and tumor-related factors have been proposed as prognostic factors (Bretheau et al. (1995) “Prognostic value of nuclear grade of renal cell carcinoma,” Cancer (Phila.), 76:2543-2549, Elson et al. (1988) “Prognostic factors for survival in patients with recurrent or metastatic renal cell carcinoma,” Cancer Res. 48:7310-7313, Motzer et al. (1999) “Survival and prognostic stratification of 670 patients with advanced renal cell carcinoma,” J. Clin. Oncol. 17:2530-2540, and Bui et al. (2001) “Prognostic factors and molecular markers for renal cell carcinoma,” Exp. Rev. Anticancer Ther. 1:565-575). Therefore, understanding how the complex interactions between multiple prognostic factors contribute to the clinical behavior of RCC is important for patient assessment, outcome prediction, and therapy planning.
Carbonic anhydrase IX (CAIX) protein, a member of the carbonic anhydrase family, is thought to play a role in the regulation of cell proliferation in response to hypoxic conditions and may be involved in oncogenesis and tumor progression (Pastorek et al. (1994) “Cloning and characterization of MN, a human tumor-associated protein with a domain homologous to carbonic anhydrase and a putative helix-loop-helix DNA binding segment,” Oncogene 9:2877-2888, Wykoff et al. (2000) “Identification of novel hypoxia dependent and independent target genes of the von Hippel-Lindau (VHL) tumour suppressor by mRNA differential expression profiling,” Oncogene 19:6297-6305, and Ivanov et al. (2001) “Expression of hypoxia-inducible cell-surface transmembrane carbonic anhydrases in human cancer,” Am. J. Pathol. 158:905-919). Previous studies using a monoclonal antibody against CAIX have shown that CAIX is induced constitutively in certain tumor types but is absent in most normal tissues with the exception of epithelial cells of the gastric mucosa (Ivanov et al. (2001) Am. J. Pathol. 158:905-919 (above), Zavada et al. (1993) “Expression of MaTu-MN protein in human tumor cultures and in clinical specimens,” Int. J. Cancer 54:268-274, Oosterwijk et al. (1986) “Monoclonal antibody G250 recognizes a determinant present in renal cell carcinoma and absent from normal kidney,” Int. J. Cancer 38:489-494, and Murakami et al. (1999) “MN/CA9 gene expression as a potential biomarker in renal cell carcinoma,” BJU Int. 83:743-747). Furthermore, previous immunobiochemical studies of malignant and benign renal tissues revealed that CAIX was also highly expressed in RCC, suggesting that CAIX expression may be a useful diagnostic biomarker (Uemura et al. (1999) “MN/CA IX/G250 as a potential target for immunotherapy of renal cell carcinomas,” Br. J. Cancer 81:741-746 and Liao et al. (1997) “Identification of the MN/CA9 protein as a reliable diagnostic biomarker of clear cell carcinoma of the kidney,” Cancer Res., 57: 2827-2831). Clinical tumor targeting studies by inter venous injection with a monoclonal antibody to CAIX have shown localization to RCC tumors in a mouse tumor model (Steffens et al. (1999) “Immunohistochemical analysis of tumor antigen saturation following injection of monoclonal antibody G250,” Anticancer Res. 1197-1200) and have been applied in clinical trials to treat metastatic RCC (Steffens et al. (1997) “Targeting of renal cell carcinoma with iodine-131-labeled chimeric monoclonal antibody G250,” J. Clin. Oncol. 15:1529-1537, Divgi et al. (1998) “Phase I/II radioimmunotherapy trial with iodine-131-labeled monoclonal antibody G250 in metastatic renal cell carcinoma,” Clin. Cancer Res. 4:2729-2739, and Steffens et al. (1999) “Phase I radioimmunotherapy of metastatic renal cell carcinoma with 131I-labeled chimeric monoclonal antibody G250,” Clin. Cancer Res. 3268s-3274s). However, prior to the present invention the relationship between CAIX expression and RCC survivorship was unknown.
Traditionally, stage, grade, and performance status have been the most useful predictors of outcome for RCC (Zisman et al. (2001) “Improved prognostication of renal cell carcinoma using an integrated staging system,” J. Clin. Oncol. 19:1649-1657). However, molecular markers can make a significant impact on the diagnosis and treatment of RCC. Tumor markers provide not only prognostic information to aid in the identification of patients at risk for recurrence or metastasis but can also facilitate the rational use of targeted therapeutic interventions as well. This concept has been demonstrated for the molecular marker, Her2/neu, and its use in the prognosis and treatment of breast cancer (Slamon et al. (2001) “Use of chemotherapy plus a monoclonal antibody against HER2 for metastatic breast cancer that overexpresses HER2,” N. Engl. J. Med. 344:783-792). Furthermore, the recent development of microarray technologies and other analytical techniques are permitting the rapid identification and validation of diagnostic and molecular markers.
Previous immunohistochemical investigations (Liao et al. (1994) “Identification of the MN antigen as a diagnostic biomarker of cervical intraepithelial squamous and glandular neoplasia and cervical carcinomas,” Am. J. Pathol. 145:598-609) have suggested that CAIX is a potential diagnostic biomarker for cervical neoplasms. An immunohistochemical study of RCC (Liao et al. (1997) Cancer Res. 57:2827-2831 (above)) also reported that CAIX was expressed in all examined RCCs, including granular, spindle, and papillary carcinomas, but not in those consisting of chromophobe histology or in benign renal lesions, including oncocytomas. Yet another study of 187 RCC reported CAIX expression in 87% of tumors by immunohistochemistry (Uemura et al. (1999) “MN/CA IX/G250 as a potential target for immunotherapy of renal cell carcinomas,” Br. J. Cancer 81:741-746). In a study of 321 primary clear cell RCC tumors, the inventors confirmed the high specificity of CAIX staining with 94% positive staining in clear cell carcinomas in the kidney. Other studies have reported that CAIX detection by reverse transcription-PCR assays in tumor specimens have a high correlation with immunohistochemistry (Uemura et al. (1999) Br. J. Cancer 81:741-746 (above) and Murakami et al. (1999) “MN/CA9 gene expression as a potential biomarker in renal cell carcinoma,” BJU Int. 83:743-747).
Targeted therapies directed at CAIX are being developed to exploit the exclusivity of CAIX expression in RCC for the treatment of metastatic disease. For example, early Phase I and II clinical trials have been addressing the feasibility of radioimmunotherapy using a monoclonal antibody against CAIX coupled to a radioisotope and have shown only minor therapeutic responses for patients with metastatic RCC (Steffens et al. (1999) Anticancer Res. 1197-1200 (above), Steffens et al. (1997) 15:1529-1537 (above), Divgi et al. (1998) Clin. Cancer Res. 4:2729-2739 (above), and Steffens et al. (1999) Clin. Cancer Res. 3268s-3274s (above)). Other therapy modalities target the immunogenicity of CAIX as a RCC tumor antigen (Vissers et al. (1999) “The renal cell carcinoma associated antigen G250 encodes a human leukocyte antigen (HLA)-A2.1-restricted epitope recognized by cytotoxic T lymphocytes,” Cancer Res. 59:5554-5559) by developing tumor-cell vaccines and dendritic cell vaccines (Uemura et al. (1994) “Vaccination with anti-idiotype antibodies mimicking a renal cell carcinoma-associated antigen induces tumor immunity,” Int. J. Cancer 58:555-561 and Tso et al. (2001) “Induction of G250-targeted and T-cell-mediated antitumor activity against renal cell carcinoma using a chimeric fusion protein consisting of G250 and granulocyte/monocyte-colony stimulating factor,” Cancer Res. 61:7925-7933). Yet another targeted approach would be to inhibit CAIX activity with chemical inhibitors. A recent study reported that a carbonic anhydrase inhibitor, acetazolamide, was able to inhibit the invasive capacity of renal cancer cells in vitro (Parkkila et al. (2000) “Carbonic anhydrase inhibitor suppresses invasion of renal cancer cells in vitro,” Proc. Natl. Acad. Sci. USA 97:2220-2224). Because CAIX is a cell surface protein unlike intracellular carbonic anhydrase isoenzymes, the design of specific chemical inhibitors of CAIX that are cell impermeable may demonstrate higher selectivity and less toxicity for suppressing renal cancer invasion.
The molecular role of CAIX in tumorigenesis is currently being elucidated, and RCC provides a unique model to study the role of hypoxia in solid tumor oncogenesis and progression. Constitutive expression of CAIX as a result of von Hippel-Lindau protein mutations (Ivanov et al. (1998) “Down-regulation of transmembrane carbonic anhydrases in renal cell carcinoma cell lines by wild-type von Hippel-Lindau transgenes,” Proc. Natl. Acad. Sci. USA 95:12596-12601) has been described for RCC. However, recent studies now indicate that expression of CAIX is regulated by the hypoxia-inducible factor 1 transcriptional complex that mediates expression of a number of genes in response to hypoxic conditions (Wykoff et al. (2000) “Hypoxia-inducible expression of tumor-associated carbonic anhydrases,” Cancer Res. 60:7075-7083). Furthermore, higher CAIX expression has been reported in perinecrotic regions of several tumor types (Ivanov et al. (2001) Am. J. Pathol. 158:905-919 (above) and Olive et al. (2001) “Carbonic anhydrase 9 as an endogenous marker for hypoxic cells in cervical cancer,” Cancer Res. 61:8924-8929). It has been postulated that cell surface carbonic anhydrases regulate acid-base balance to optimize conditions in the tumor invasiveness (Ivanov et al. (2001) Am. J. Pathol. 158:905-919 (above)). Acidification of the extracellular matrix is known to induce expression of angiogenic factors (Shi et al. (1999) “Constitutive and inducible interleukin 8 expression by hypoxia and acidosis renders human pancreatic cancer cells more tumorigenic and metastatic,” Clin. Cancer Res. 5:3711-3721) and may inhibit cellular immunity (Giatromanolaki et al. (2001) “Expression of hypoxiainducible carbonic anhydrase-9 relates to angiogenic pathways and independently to poor outcome in non-small cell lung cancer,” Cancer Res. 61:7992-7998), which additionally promotes tumor aggressiveness. In addition, there is some evidence for the association of CAIX with loss of contact inhibition and anchorage dependence of cancer cells (Parkkila et al. (2000) Proc. Natl. Acad. Sci. USA 97:2220-2224 (above)).
From the foregoing, it is apparent that methods of correlating CAIX expression with RCC survivorship are desirable. More specifically, it is desirable to predict clinical outcome and/or to identify high-risk patients in need of adjuvant immunotherapy, CAIX-targeted therapies, or other methods of treatment based, at least in part, on CAIX expression levels. These and a variety of additional features of the present invention will become evident upon complete review of the following disclosure.